title: Combining mTOR inhibition with radiation improves antitumor activity in bladder cancer cells in vitro and in vivo: a novel strategy for treatment.
abstract: OBJECTIVE
  Radiation therapy for invasive bladder cancer allows for organ preservation but toxicity and local control remain problematic. As such, improving efficacy of treatment requires radiosensitization of tumor cells. The aim of study is to investigate if the mammalian Target of Rapamycin (mTOR), a downstream kinase of the phosphatidylinositol 3-kinase (PI3K)/AKT survival pathway, may be a target for radiation sensitization.
  
  
  METHODS
  Clonogenic assays were performed using 6 bladder cancer cell lines (UM-UC3, UM-UC5, UM-UC6, KU7, 253J-BV, and 253-JP) in order to examine the effects of ionizing radiation (IR) alone and in combination with RAD001, an mTOR inhibitor. Cell cycle analysis was performed using flow cytometry. In vivo, athymic mice were subcutaneously injected with 2 bladder cancer cell lines. Treatment response with RAD001 (1.5 mg/kg, daily), fractionated IR (total 9Gy = 3Gy×3), and combination of RAD001 and IR was followed over 4 weeks. Tumor weight was measured at experimental endpoint.
  
  
  RESULTS
  Clonogenic assays revealed that in all bladder cell lines tested, an additive effect was observed in the combined treatment when compared to either treatment alone. Our data indicates that this effect is due to arrest in both G1 and G2 phases of cell cycle when treatments are combined. Furthermore, our data show that this arrest is primarily regulated by changes in levels of cyclin D1, p27 and p21 following treatments. In vivo, a significant decrease in tumor weight was observed in the combined treatment compared to either treatment alone or control.
  
  
  CONCLUSIONS
  Altering cell cycle by inhibiting the mTOR signaling pathway in combination with radiation have favorable outcomes and is a promising therapeutic modality for bladder cancer.
authors.affiliation: McGill Urologic Oncology Research, Division of Urology, McGill University, Montreal, Quebec, Canada.
authors.forename: Roland
authors.initials: R
authors.lastname: Nassim
authors.forename: Jose Joao
authors.initials: JJ
authors.lastname: Mansure
authors.forename: Simone
authors.initials: S
authors.lastname: Chevalier
authors.forename: Fabio
authors.initials: F
authors.lastname: Cury
authors.forename: Wassim
authors.initials: W
authors.lastname: Kassouf
mesh_terms: D045744:Cell Line, Tumor; D016193:G1 Phase / Q000528:radiation effects; D016195:G2 Phase / Q000528:radiation effects; D006801:Humans; D066298:In Vitro Techniques; D011836:Radiation Tolerance; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors*; D001749:Urinary Bladder Neoplasms / Q000473:pathology / Q000532:radiotherapy*
chemical_list: C546842:MTOR protein, human; D058570:TOR Serine-Threonine Kinases